Aberrant hyperactivation of KRAS plays a prominent role in tumor initiation and progression in a broad spectrum of human cancers. The incidence of KRAS mutations is especially high in colorectal malignancies, where it occurs at a frequency of >40%. We hypothesize that dual targeting of CDK4 and MEK will offer clear therapeutic benefit for the subpopulation of colorectal patients whose tumors are addicted to MEK signaling. The overarching goal of this proposal is to generate compelling preclinical support to inform rationally designed clinical trials of CDK4/MEK inhibitor-based combination therapies for metastatic colorectal cancer patients. Critical to the success of this project will be incorporatio of preclinical animal models that are predictive of clinical disease. Therefore, we propose to study a panel of primary human xenograft models established from specimens procured from surgeries carried out at our Institution in an attempt to recapitulate the heterogeneity encountered in the CRC patient population. All models will be molecularly profiled for genomic aberrations implicated in colorectal cancer progression, including analysis of KRAS and RB. Using the clinical candidate palbociclib (PD0332991) and clinically approved trametinib, which are highly selective for CDK4/6 and MEK, respectively, we propose to carry out pharmacologic profiling of our entire cohort of RB+ xenografts to identify the subpopulation of models that are responsive to both single agents. Focusing on this subset of models judged to have the highest likelihood of responding to a CDK4/MEK co-targeting strategy, we propose to employ a molecular imaging approach to design the optimal combination regimen for these two agents. Our primary human CRC models will be transduced with Luc2-IRES-mCherry lentivirus to enable bioluminescent imaging of liver lesions resulting from disseminated disease after orthotopic implantation into the cecum. Multiple doses and schedules will be evaluated as we compare therapeutic outcome of concurrent versus sequential dosing strategies. We believe this to be the first study undertaken to optimize co-targeting of CDK4 and MEK to treat colorectal cancer on the basis of bioluminescent imaging of metastatic lesions originating and proliferating directly in the mouse liver.